This application seeks support for a midcareer investigator to mentor appropriate candidates in mouse pathobiology research. Dr. David Huso is an independent researcher and pathologist in the Division of Comparative Medicine at JHU. He has increasingly focused on studies of genetically engineered mice and uses them in his research to understand HIV neuropathogenesis and persistence. As a pathology faculty member of a combined training program in Laboratory Animal Medicine and Comparative Pathology at JHU, Dr. Huso has assisted in training over two dozen veterinary fellows in these specialities over the past ten years. In addition, at least two veterinary students, as well as preveterinary students and veterinary assistants each year choose to spend preceptorships in the division. These fellows and students provide a pool of talented individuals to recruit and mentor in mouse pathobiology research. JHU provides a rich environment for mouse pathobiology research. Currently, JHU supports core transgenic and knockout laboratories with close ties to the Division of Comparative Medicine. A growing emphasis within the division is on phenotype analysis and gene function studies of genetically engineered mice. Plans for the university include marked expansion of facilities to support studies of mutant mice. There are 437 protocols at JHU which use mice and an ever-expanding annual use of more than 70,000 mice. This creates a high level of enthusiasm for mouse pathobiology research at JHU. Dr. Huso has NIH R01 support to use mice transgenic for human cytokines to create a primary neuronal-glial culture system to study HIV and the ramification and activation of microglia. Clearance of cellular reservoirs that harbor HIV during highly active antiretroviral therapy treatment is mainly dependent on the particular turnover rate of the reservoir target cell population. Most of the HIV-1-infected cells in the brain are macrophages and microglia. Ramified, tissue microglia in the central nervous system represent one of the most stable cell types in the body. Virus could persist in these quiescent cells for years or perhaps even decades before the cells turn over. However, since poorly characterized cellular interactions and unknown soluble factors maintain microglial ramification in the brain and because glial are inaccessible, the biology of ramified microglia is still poorly understood. The proposed studies could give new insights into HIV persistence, microglia infection, and neuronal injury. This revised K26 research plan proposes to extend these R01 studies by using mice transgenic for the HIV co-receptors, human CCR5 and CD4: 1) to develop an in vivo model for HIV neuropathogenesis and persistence in transgenic mice; 2) to compare the entry of HIV-1 in ramified and activated microglia; and 3) to determine the effects of CD4 or CCR5 receptor binding on superoxide production in microglia.